The objective of the Program is to study the molecular regulation of pattern formation in the developing vertebrate limb, and to begin to apply this basic knowledge to understanding the nature of human genetic disorders affecting limb patterning. In Project 1, the role that apoptosis plays in maintenance of the activity of the apical ectodermal ridge (AER) that directs limb outgrowth and patterning will be investigated, as will the related hypothesis that the homeobox- containing gene Msx-2 and BMPs are in a regulatory network that controls programmed cell death in the AER. The hypothesis that the homeobox- containing gene Cux-, the vertebrate homolog of the Drosophila Cut gene, AER. The hypothesis that the homeobox-containing Cux, the vertebrate homolog of the Drosophila Cut gene, plays a critical role in regulation the onset of the segmentation process that generates joints between the skeletal elements of the limb will also be studied. In Project 2, the roles and relationships among the homeobox-containing gene Dlx-5, and the growth factors IGF-1, TGF-alpha, and FGF-10 in the establishment of the presumptive limb regions of the lateral plate, and the initial formation of the limb from these regions will be investigated. In Project 3, the transcriptional mechanisms that regulate the expression of Msx-2 in the developing vertebrate limb will be studied. Particular emphasis will be on identification of the cis-regulatory elements and trans-acting factors that control the independently-regulated spatially- specific domains of Msx-2 expression and transacting factors that control the independently-regulated spatially-specific domains of Msx-2 expression in the AER and mesoderm. In Project 4, genetic linkage analyses and positioning cloning strategies will be used to map the loci of and identify the genes responsible for the human genetic defects Split Hand-Split Foot-4 malformations and Acheiropodia, which are genetic disorders that result from impairments in developmental patterning of the limbs. A Core unit will supply the Projects with amelic and polydactylous mutant chick embryos that will be used in the analysis of the roles of regulatory genes and signaling molecules in patterning. The integrated goal of the Program is to study the nature, regulation, interactions, and mechanisms of action of some of the genes and signaling molecules involved in the formation, outgrowth, and patterning of the vertebrate limb, and to begin to apply this basic knowledge the nature of human genetic disorders affecting limb patterning.